Therapeutic silencing of mTOR by systemically administered siRNA-loaded neutral liposomal nanoparticles inhibits DMBA-induced mammary carcinogenesis.

BACKGROUND: Mammary carcinogenesis possesses great challenges due to the lack of effectiveness of the multiple therapeutic options available. Gene therapy-based cancer treatment strategy provides more targeting accuracy, fewer side effects, and higher therapeutic efficiency. Downregulation of the oncogene mTOR by mTOR-siRNA is an encouraging approach to reduce cancer progression. However, its employment as means of therapeutic strategy has been restricted due to the unavailability of a suitable delivery system. METHODS: A suitable nanocarrier system made up of 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) has been developed to prevent degradation and for proficient delivery of siRNA. This was followed by in vitro and in vivo anti-breast cancer efficiency analysis of the mTOR siRNA-loaded neutral liposomal formulation (NL-mTOR-siRNA). RESULTS: In our experiment, a profound reduction in MCF-7 cell growth, proliferation and invasion was ascertained following extensive downregulation of mTOR expression. NL-mTOR-siRNA suppressed tumour growth and restored morphological alterations of DMBA-induced breast cancer. In addition, neutral liposome enhanced accumulation of siRNA in mammary cancer tissues facilitating its deep cytosolic distribution within the tumour, which allows apoptosis thereby facilitating its anti-tumour potential. CONCLUSION: Hence, the current study highlighted the augmented ground for therapies aiming toward cancerous cells to diminish mTOR expression by RNAi in managing mammary carcinoma.

Structure Based Multitargeted Molecular Docking Analysis of Selected Furanocoumarins against Breast Cancer.

Breast cancer is one of the biggest global dilemmas and its current therapy is to target the hormone receptors by the use of partial agonists/antagonists. Potent drugs for breast cancer treatment are Tamoxifen, Trastuzumab, Paclitaxel, etc. which show adverse effects and resistance in patients. The aim of the study has been on certain phytochemicals which has potent actions on ERα, PR, EGFR and mTOR inhibition. The current study is performed by the use of molecular docking as protein-ligand interactions play a vital role in drug design. The 3D structures of ERα, PR, EGFR and mTOR were obtained from the protein data bank and docked with 23 3D PubChem structures of furanocoumarin compounds using FlexX. Drug-likeness property was checked by applying the Lipinski's rule of five on the furanocoumarins to evaluate anti-breast cancer activity. Antagonist and inhibition assay of ERα, EGFR and mTOR respectively has been performed using appropriate in-vitro techniques. The results confirm that Xanthotoxol has the best docking score for breast cancer followed by Bergapten, Angelicin, Psoralen and Isoimperatorin. Further, the in-vitro results also validate the molecular docking analysis. This study suggests that the selected furanocoumarins can be further investigated and evaluated for breast cancer treatment and management strategies.

Bergapten inhibits liver carcinogenesis by modulating LXR/PI3K/Akt and IDOL/LDLR pathways.

Oxysterol receptors LXRs (α and ß) are recently reported to be one of the novel and potential therapeutic targets in reducing cell proliferation and tumor growth in different system model. Activation of LXRs is correlated with modification of PI3K/Akt pathway. LXRs are also found to play a critical role in maintaining lipid homeostatais by regulating ABCA1, IDOL, SREBP1, LDLR and also certain lipogenic genes such as FASN and SCD1. In the present study a potential furanocoumarin, Bergapten (BeG) has been evaluated for its anticancer property on Hepatocellular Carcinoma (HCC) on LXR axis. The molecular docking analysis was carried out for BeG on LXR (α & ß) using Maestro tool and compared with reference ligands. This was followed by in vitro (HepG2 cell lines) and in vivo (on NDEA induced HCC in Wistar albino rats) anticancer evaluation of BeG. The docking results revealed polar and hydrophobic interactions of BeG with LXR (α,ß). The in vitro studies revealed the potential of BeG in lowering the accumulation of lipid droplets in HepG2 cells which was correlated with increase in LXR (α,ß) protein expressions. Furthermore, the in vivo studies demonstrated the potential of BeG in ameliorating the cancer induced alterations in body weight, liver weight and significant restoration of the changes in mRNA and protein expressions of LXR(α,ß), ABCA1, IDOL, SREBP1 and LDLR. BeG also modulated the expressions of PI3K, Akt and certain lipogenic genes like FASN and SCD1 and reduced the lipid droplets level in liver cancer cells. These results provide evidence and validates the critical role of BeG in maintaining the lipid homeostasis and justifies its anticancer potential against NDEA-induced HCC.

mTORC1 activation in B cells confers impairment of marginal zone microarchitecture by exaggerating cathepsin activity.

Mammalian target of rapamycin complex 1 (mTORC1) is a key regulator of cell metabolism and lymphocyte proliferation. It is inhibited by the tuberous sclerosis complex (TSC), a heterodimer of TSC1 and TSC2. Deletion of either gene results in robust activation of mTORC1. Mature B cells reside in the spleen at two major anatomical locations, the marginal zone (MZ) and follicles. The MZ constitutes the first line of humoral response against blood-borne pathogens and undergoes atrophy in chronic inflammation. In previous work, we showed that mice deleted for TSC1 in their B cells (TSC1BKO ) have almost no MZ B cells, whereas follicular B cells are minimally affected. To explore potential underlying mechanisms for MZ B-cell loss, we have analysed the spleen MZ architecture of TSC1BKO mice and found it to be severely impaired. Examination of lymphotoxins (LTα and LTß) and lymphotoxin receptor (LTßR) expression indicated that LTßR levels in spleen stroma were reduced by TSC1 deletion in the B cells. Furthermore, LTα transcripts in B cells were reduced. Because LTßR is sensitive to proteolysis, we analysed cathepsin activity in TSC1BKO . A higher cathepsin activity, particularly of cathepsin B, was observed, which was reduced by mTORC1 inhibition with rapamycin in vivo. Remarkably, in vivo administration of a pan-cathepsin inhibitor restored LTßR expression, LTα mRNA levels and the MZ architecture. Our data identify a novel connection, although not elucidated at the molecular level, between mTORC1 and cathepsin activity in a manner relevant to MZ dynamics.

Taxifolin Inhibits 7,12-Dimethylbenz(a)anthracene-induced Breast Carcinogenesis by Regulating AhR/CYP1A1 Signaling Pathway.

BACKGROUND: Breast cancer (BC), because of its invasive characteristics, is one of the most common and deadliest cancers among the female population around the world. Research has demonstrated that AhR signaling also plays a vital role in BC initiation and development as well. Therefore, blocking this pathway to natural interferences paves a new channel for the prevention of BC. Several natural compounds such as flavonoids possess the anticancer activities against different cancers. OBJECTIVE: The present study has been designed to estimate the chemotherapeutic potential of taxifolin (TAX) against 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary carcinoma in Sprague-Dawley rats. MATERIALS AND METHODS: Initially, the molecular docking analysis of AhR and cytochrome P450s (CYPs) (CYP1A1 and CYP1B1) was performed using MAESTRO tool, in an attempt to rationalize the activity of TAX, based on their CYP1-binding potential. The in vitro CYP1A1 activity was determined by luciferase assay with CYP1A1 substrate luciferin CEE. The in vivo analysis was performed by administrating TAX at 10, 20, 40 mg/kg BW for 28 days intragastrically in DMBA induced (25 mg/animal dose) at 55 days of age Sprague-Dawley (SD) rats. BC initiates after 90 days of tumor induction phase. The molecular mechanism of TAX on Ahr and CYPs was also examined through the mRNA and protein expressions using reverse transcription-quantitative polymerase chain reaction and Western blotting analysis. RESULTS: Furthermore, TAX altered the energy regulation on DMBA-induced BC in SD rats by considerably restoring the cancer-induced modulations in tumor growth. Our results showed that TAX reduced the expressions of CYP1A1 and CYP1B1 in DMBA-induced mammary carcinoma by downregulating the AhR signaling pathway. CONCLUSION: This study revealed that TAX might be able to act as a chemotherapeutic agent against CYP1A1- and CYP1B1-mediated cancer and the inhibition of the DMBA-induced mammary carcinogenesis in a rat model.Abbreviations used: CYPs: Cytochrome P450s; PAH: polycyclic aromatic hydrocarbons; HRP- Horseradish peroxidase; BSA: Bovine serum albumin; DTTP: Deoxythymidine Triphosphate (nucleotide); RT-qPCR: Real Time quantitative polymerase chain reaction; CADD: Computer Aided Drug Drafting.

Daphnetin inhibits TNF-α and VEGF-induced angiogenesis through inhibition of the IKKs/IκBα/NF-κB, Src/FAK/ERK1/2 and Akt signalling pathways.

Coumarins, identified as plant secondary metabolites possess diverse biological activities including anti-angiogenic properties. Daphnetin (DAP), a plant derived dihydroxylated derivative of coumarin has shown significant pharmacological properties such as anticancer, anti-arthritic and anti-inflammatory. The present study was performed to investigate the anti-angiogenic potential of DAP, focusing on the mechanism of action. The in vivo anti-angiogenic potential of DAP was evaluated by vascular endothelial growth factor (VEGF)-induced rat aortic ring (RAR) assay and chick chorioallantoic membrane (CAM) assay. For in vitro evaluation, wounding migration, transwell invasion, tube formation and apoptosis assays were performed on VEGF (8 ng/mL)-induced human umbilical vein endothelial cells (HUVECs). The cellular mechanism of DAP was examined on TNFα (10 ng/mL) and VEGF-induced HUVECs by extracting the mRNA and protein levels using RT-qPCR and western blotting. Our data demonstrated that DAP inhibited the in vivo angiogenesis in the RAR and CAM assay. DAP also inhibited the different steps of angiogenesis, such as migration, invasion, and tube formation in HUVECs. DAP inhibited nuclear factor-κB signalling together including TNF-α induced IκBα degradation; phosphorylation of IκB kinase (IKKα/ß) and translocation of the NF-κB-p65 protein. Furthermore, western blotting revealed that DAP significantly down-regulated the VEGF-induced signalling such as c-Src, FAK, ERK1/2 and the related phosphorylation of protein kinase B (Akt) and VEGFR2 expressions. DAP reduced the elevated mRNA expression of iNOS, MMP2 and also, induced apoptosis in VEGF-stimulated HUVECs by the caspase-3 dependent pathway. Taken together, this study reveals that DAP may have novel prospective as a new multi-targeted medication for the anti-angiogenesis and cancer therapy.

Bronchodilatory and mast cell stabilising activity of Cressa cretica L.: evaluation through in vivo and in vitro experimental models.

OBJECTIVE: To evaluate the effect of ethylacetate fraction (Fr-Et) and methanolic fraction (Fr-Me) obtained from Cressa cretica L.(C. cretica) L. on experimental models for bronchodilatory activity and mast cell stabilising activity. METHODS: The effect of Fr-Et and Fr-Me were studied on acetylcholine and histamine aerosol-induced broncospasm using guinea pigs as experimental animals. Also, the effects of these fractions were evaluated on the isolated guinea pig tracheal preparations. Besides this mast cell degranulation effect was assessed using egg albumin and compound 48/80 on rat peritoneal mast cells. RESULTS: Significant increase in preconvulsion time was observed due to pretreatment with the fractions when guinea pigs were exposed to histamine and acetylcholine aerosol. Fr-Et and Fr-Me significantly increased the preconvulsion in a dose depended manner that suggestive of bronchodilating activity. Fr-Et and Fr-Me exhibited a significant concentration dependant relaxant effect on guinea pig trachea pre-contracted with CCh, K(+) and histamine. The results revealed that Fr-Et to be more potent than Fr-Me in relaxing histamine and K(+) and calcium induced contraction than CCh induced contractions. Studies on the fractions in protecting mast cell degranulation, which were elicited by the egg albumin as well as synthetic compound 48/80 revealed both the fractions significantly protect the mast cell degranulation, which release mediators such as histamine and proinflammatory cytokines through various stimuli in a dose depended manner. CONCLUSIONS: Thus our study established the bronchodilator activity, and mast cell stabilizing activity which are important mediators that provoke or sustain in asthma.

Effect of Dendrophthoe falcata (L.f.) Ettingsh on female reproductive system in Wistar rats: a focus on antifertility efficacy.

BACKGROUND: This study was designed to assess the effect of one traditionally used antifertility plant, Dendrophthoe falcata (L.f.) Ettingsh (Loranthaceae), on female reproductive system, fertility and safety, by oral administration to adult female Wistar rats. STUDY DESIGN: After the oral acute toxicity study, the hydroalcoholic extract of the aerial parts was administered at three doses. Treatment started at weaning and continued until Day 0 postpartum. RESULTS: The LD(50) value was found to be 4.55 g/kg body weight. Vaginal opening occurred earlier in treated females when compared with controls. Analysis of vaginal smears revealed that all animals were cycling, although the length of the diestrus was longer in treated groups. In postcoital testing, the extract was found to be more effective in causing significant anti-implantation activity and reduction in the number of litters born. The extract also exhibited weak estrogenic activity when given alone, and when given along with ethinyl estradiol, it exhibited slight antiestrogenic activity in immature ovariectomized rats. CONCLUSION: All observations suggest that the extract has antifertility effect and is safe at effective doses employed in the study.